Oligonucleotide therapeutic strategies rely on delivering the hydrophillic, highly charged and anionic nucleic acid molecule into the cytoplasm by crossing the hydrophobic phospholipid bilayer that constitutes the cell membrane. Hydrophobic conjugates of oligonucleotides have been investigated for their ability to enhance cellular uptake. For example, cholesterol conjugates may be capable of binding to lipoproteins, and more specifically IDL and LDL particles. The particles contain ApoE and ApoB-100 lipoproteins, which target LDL cell surface receptors and, therefore, can be internalized via lipoprotein mediated endocytosis. These receptors are found mainly in the liver, but also in adipose tissue, heart and skeletal muscle. Additionally, pendant lipophillic groups may aid in “anchoring” their conjugates to the cell membrane by intercalating into the phospholipid bilayer and therefore aiding passive internalization of the oligonucleotide.
Such lipophilic molecules or substitutents must be formulated with, or attached to, the polynucleotide so as to preserve the functional and structural integrity of the polynucleotide for its desired therapeutic action. For example, therapeutic polynucleotides may function through interactions with large and/or intricate ribonucleoprotein (RNP) complex(es), and/or may target RNA components of these complexes. Lipophilic modifications should therefore not impair the accessibility of the polynucleotide for interactions with cellular factors and molecular targets.
MicroRNAs (miRs) have been implicated in a number of biological processes including regulation of developmental timing, apoptosis, fat metabolism, and hematopoietic cell differentiation among others, and therefore represent a relatively new class of therapeutic targets. miRs are small, non-protein coding RNAs of about 18 to about 25 nucleotides in length, and act as repressors of target mRNAs by promoting their degradation, when their sequences are perfectly complementary, or by inhibiting translation, when their sequences contain mismatches. The mature miRNA strand is incorporated into the RNA-induced silencing complex (RISC), where it associates with its target RNAs by base-pair complementarity.
Compositions and methods for efficiently delivering therapeutic polynucleotides, including miRs and miR inhibitors, to cells without substantially or significantly impairing interactions with the molecular target(s) are needed.